The aims of the research in the past year was to quantify integrin alpha(V)beta(3) receptor expression on tumor-induced angiogenic blood vessels with In-111-labeled antagonists during a course of therapy, and to investigate a co-relationship between the level of the receptor expression and therapeutic response reflected in the changes in tumor size. Our preliminary studies demonstrated that In-111-labeled second generation peptidomimetic antagonist and In-111-labeled dimeric cyclic RGD peptide were good molecular targeting agents for alpha(V)beta(3) receptor. Among these In-111 labeled reagents, In-111 labeled dimeric cyclic RGD peptide was most suitable for quantifying the changes in the level of the receptor. Methods: Groups of nude mice (n = 5-10 per time point) implanted s.c. with A431/K5 tumor cells, expressing mesothelin and not alpha(V)beta(3), were treated with taxol alone i.p. at 50 mg/kg on day 0, mesothelin-specific immunotoxin SS1P alone i.v. at 0.2 mg/kg on days 0 and 2, or the two agents together; taxol (day 0), and SS1P (days 1 and 3). The tumor size was 100 cubic mm on day 0 and was measured daily. For the assessment of therapeutic response, the mice received i.v. In-111-labeled dimeric cyclic RGD peptide (2.0 micro-Ci/<0.1 micro-g) in 0.2 ml of PBS containing 1% BSA on days 1 and 4 for the taxol alone groups and on days 1 and 3 for the immunotoxin alone groups, and were euthanized at 1 or 2 hr post-injection for biodistribution. For the combination therapy groups, the mice were injected i.v. with the radiolabel on days 2 and 4, and were euthanized at 1 or 2 hr post-injection of the radiolabel. Results: The combination therapy was synergistic, using doses at which either agent alone causes stabilization of tumor growth as reported previously (Clin Cancer Res 2006;12:4695). The biodistribution of the radiolabel in blood and major organs was similar between the control and the drug treated animals. However, compared to the control, the tumor uptake (% ID/g) of the radiolabel decreased by 25% on day 1 but increased by 33% on day 4 for the taxol treated animals. Comparatively, the uptake increased by 11% on day 1 and by 39% on day 3 for the immunotoxin treated animals. The combined therapy decreased the uptake (% ID/g) by 8.5% on day 2 but increased the uptake by 25% on day 4. Since the taxol or immunotoxin treatment stopped the tumor growth and the tumor sizes remained unchanged for the duration of this experiment, the negative changes in tumor uptake (% ID/g) appear to indicate that taxol treatment down-regulated the receptor expression and thus, reduced the total level of the receptors on angiogenic vessels on day 1. However, the receptor level (%ID/g) increased on day 4, indicating that the receptor expression became up-regulated to induce angiogenesis for the re-growth of tumors. In contrast, immunotoxin treatment increased the receptor level for a 4 day period, indicating that immunotoxin treatment up-regulated the receptor expression. The changes in the tumor uptake by the combination therapy are the results of the combined effects of the drugs on the receptor expression. Conclusions: This study demonstrated that In-111 labeled dimeric cyclic RGD peptide is a good molecular targeting agent for alpha(V)beta(3) receptors and that the changes in the level of the alpha(V)beta(3) receptors (%ID/g tumor) expressed on neovasculature could be used as a sensitive surrogate marker for therapeutic response. The opposite direction of the changes in the receptor level suggests that the therapeutic effects of these two drugs were induced by different pathways of action. The direction and the magnitude of the changes in the receptor expression may be a sensitive predictive marker for therapeutic response of drugs.